The Impact of Selection with Diflubenzuron, a Chitin Synthesis Inhibitor, on the Fitness of Two Brazilian Aedes aegypti Field Populations

Abstract

Several Aedes aegypti field populations are resistant to neurotoxic insecticides, mainly organophoshates and pyrethroids, which are extensively used as larvicides and adulticides, respectively. Diflubenzuron (DFB), a chitin synthesis inhibitor (CSI), was recently approved for use in drinking water, and is presently employed in Brazil for Ae. aegypti control, against populations resistant to the organophosphate temephos. However, tests of DFB efficacy against field Ae. aegypti populations are lacking. In addition, information regarding the dynamics of CSI resistance, and characterization of any potential fitness effects that may arise in conjunction with resistance are essential for new Ae. aegypti control strategies. Here, the efficacy of DFB was evaluated for two Brazilian Ae. aegypti populations known to be resistant to both temephos and the pyrethroid deltamethrin. Laboratory selection for DFB resistance was then performed over six or seven generations, using a fixed dose of insecticide that inhibited 80% of adult emergence in the first generation. The selection process was stopped when adult emergence in the diflubenzuron-treated groups was equivalent to that of the control groups, kept without insecticide. Diflubenzuron was effective against the two Ae. aegypti field populations evaluated, regardless of their resistance level to neurotoxic insecticides. However, only a few generations of DFB selection were sufficient to change the susceptible status of both populations to this compound. Several aspects of mosquito biology were affected in both selected populations, indicating that diflubenzuron resistance acquisition is associated with a fitness cost. We believe that these results can significantly contribute to the design of control strategies involving the use of insect growth regulators.

Fig 1. Stage-specific mortality observed for Ae. aegypti mosquitoes in response to treatment with different concentrations of the insecticide DFB.

Mosquitoes from two field-derived Brazilian populations; Boa Vista (BVT) and Aparecida de Goiânia (APG) were compared prior to selection with DFB (Panel A), and then again after the BVT and APG populations had been reared for several generations in the absence (cont) or presence (dfb) of diflubenzuron (Panel B). Note, that in the two rightmost graphs higher concentrations of DFB were used. The selection process lasted for six generations for BVT, and seven for APG. The Rockefeller strain was used as a susceptible control both before (Rock A), and after selection (Rock B).

Fig 2. The sex ratio of mosquitoes both pre- (Panel A) and post-selection, (Panel B) showing the proportion of male (blue) and female (pink) adults in each group.

The value at the bottom of each bar represents the total number of adults that eclosed in that particular treatment. Diflubenzuron-selected (dfb) and control (cont) strains are shown for the Boa Vista (BVT) and Aparecida de Goiânia (APG) populations.

Fig 3. The evolution of DFB resistance in response to successive generations of exposure for BVT (A) and APG (B) mosquitoes, as indicated by an increase in the percentage of mosquitoes surviving to adulthood.

The control groups ("cont", in blue) were kept in the laboratory for the same number of generations, and reared without DFB. Curves with the same colors represent replicas.

Fig 4. The effect of DFB on adult emergence in the Rockefeller strain (Rock B), control groups (Panel A) and groups selected with DFB (Panel B), during the bioassays conducted post-selection.

Curves depict an analysis of non-linear regression (R² > 0.9).

Fig 5. Daily mortality of the Rockefeller strain (black circles), control (blue circles) and selected (red circles) groups of BVT and APG mosquitoes.

Panels A and B depict male longevity, while panels C and D show female longevity. All curves depict an analysis of non-linear regression (R² > 0.9).

Fig 6. Rate of bloodmeal acceptance for females from the Rockefeller strain (Rock), selected (dfb), or control (cont) groups from Boa Vista (BVT) and Aparecida de Goiânia (APG).

Different letters above the columns indicate significant differences among groups (χ2; P < 0.05).

Fig 7. The amount of blood ingested by Rockefeller (Rock) females, and DFB-selected (dfb) or control (cont) mosquitoes from Boa Vista (BVT) and Aparecida de Goiânia (APG).

Each box-plot displays the median, the interquartile range and the minimum and maximum amount of blood ingested by each group. Distinct letters indicate significant differences among groups (t test; P < 0.05).

Fig 8. Number of eggs laid by Rockefeller (Rock) females, and selected (dfb) or control (cont) groups of Boa Vista (BVT) and Aparecida de Goiânia (APG) mosquitoes.

Box-plots and letters above them are as for Fig 7. All groups were compared pairwise except for control and selected groups from each tested population.

Fig 9. Male mating efficiency for Rockefeller (Rock), and selected (dfb) or control (cont) groups of Boa Vista (BVT) and Aparecida de Goiânia (APG) mosquitoes.

For each group, one male was placed in contact with three females for three days. The results are expressed as the percentage of males able to inseminate one, two or three females.